Technical Abstract:
Microarrays provide a powerful tool to examine genome wide patterns of differential transcription. We are using microarrays to identify Fusarium verticillioides' structural and regulatory genes involved in the biosynthesis of fungal toxins, virulence factors, and other elements involved in plant pathogenesis. In collaboration with The Institute for Genomics Research (TIGR), we developed a NimbleGen Systems, Inc. (Madison, WI) microarray chip containing 15,844 probe sets that represent over 11,126 different genes. Each probe set consists of up to twelve 24-mer oligonucleotides. The gene sequences represent approximately 81% of the coding capacity of F. verticillioides and were generated from analysis of over 87,000 Expressed Sequence Tags (ESTs). The major goal of this research is to develop new strategies to limit the negative impact Fusarium have on crops (e.g. maize) due to disease and contamination by mycotoxins. The main set of microarray data described in this paper was generated from cultures of wild-type F. verticillioides strain M-3125 in the liquid fumonisin production medium, GYAM, for 12, 24, 48, 72, 96 and 120 hrs. Mycelium was harvested from duplicate cultures at each time point and RNA was extracted and analyzed for quality. This presentation will describe three approaches we have taken to examine the data generated from this experiment. The goal of the first approach was to identify genes that were transcriptionally regulated in a manner similar to fumonisin biosynthetic genes. Transcriptional co-regulation has served to identify genes involved in the same biological process in other systems. In this manner, we hope to identify additional genes that may affect fumonisin biosynthesis. The second approach involves using the microarray data to identify genes that are co-regulated with genes that may be required in the synthesis of other secondary metabolites. We present evidence for three clusters of co-regulated genes that includes a polyketide synthase (PKSs). One of these PKSs has been shown to be involved in the synthesis of the mycotoxin fusarin. The third approach examines the specific differential regulation of FUM21 alternative splice form (ASF) transcripts. FUM21 is a narrow-domain transcriptional activator of fumonisin gene transcription. These results have provided unique insight as to how FUM21 regulates fumonisin biosynthesis. Taken together, these different approaches demonstrate the power of microarray technology to provide a wealth of information on a large number of different biological processes.